Implementation of a workflow for the detection and validation of low level somatic BRCA1/BRCA2 variants for the clinical management of high grade serous ovarian cancer patients


Ovarian cancer is the sixth most common cause of cancer in women and is estimated to cause 7,400 new cases and 4,200 deaths on an annual basis in the UK. Both BRCA1 and BRCA2 are amongst the most frequently mutated genes in high-grade ovarian serous carcinoma, and account for a total of 20% of cases; with germline mutation rate estimated at 15%, while somatic mutations account for 5%.    Because BRCA genes play a key role in the homologous recombination DNA repair pathway, an improved prognosis has been demonstrated in patients carrying BRCA alterations who were treated with an inhibitor of PARP-1 protein that plays a central role in single-stranded DNA break repair, as this causes synthetic lethality. This has led to the approval of PARP-1 inhibitor, olaparib, for use in ovarian cancer patients carrying germline or somatic mutations in either of the BRCA genes.    Although investigation of germline variants using next generation sequencing technology is well established, patients with somatic mutations cannot be reliably screened using the same methodology, since significant DNA damage occurs during routine preservation of tumour tissues using formalin fixation. Thus, most patients with somatic alterations cannot benefit from PARP inhibitor therapy. To address this, we have evaluated two amplicon-based enrichment assays, which due to the clonal nature of PCR, possess the ability to work with low yield fragmented DNA typically extracted from formalin fixed tissues. Preliminary findings show encouraging results which demonstrate the ability of such technology in detecting low level variants in formalin fixed tissues.